The long-term objective of this application is to study the role of calcium in phototransduction. Specifically, the aims of this proposal are: 1. Establish the effectiveness of controlling the concentration of cytoplasmic calcium in retinal rods. 2. Study the effects of cytoplasmic calcium at different concentrations on the dark current, the photocurrent kinetics and light-sensitivity. 3. Determination of the specific effects of cytoplasmic calcium on guanylate cyclase in detached rod outer segments. Experimental manipulations that change the concentration of cytoplasmic calcium in rods affect the amplitude of the dark current, the kinetics of the photoresponse and its light-sensitivity, yet, how calcium exerts these effects is not known. Much of the recent advance has been in the characterization of the light- sensitive channels and the mechanism that by which light activates the phosphodiesterase. Considerably less attention has been given to the processes that maintain the dark current and determine the kinetics of the turn off after a flash of light. In addition, the mechanism that allow rods to change their sensitivity (adaptation) is completely unknown. The present proposal will examine the role of cytoplasmic calcium in these processes. To achieve the goals of the present proposal I will first establish experimental control of the cytoplasmic calcium concentration. Controlling the cytoplasmic calcium of detached rod outer segments will then allow me to study the effects of calcium on the dark current and the photoresponse. After characterizing the effects of calcium on photoreceptor function I will attempt to understand the mechanism by which calcium exerts its effects. Specifically, I will test the hypothesis that calcium exerts its effects on phototransduction by regulating the activity of guanylate cyclase. I will test this hypothesis by studying its effects in the dark and in light under conditions which inhibit the phosphodiesterase.